Roughly one third of today's FDA-approved drugs act on G-protein coupled receptors (GPCRs), collectively affecting hundreds of millions of people worldwide. These medications were developed based on a limited signaling paradigm, by which stimulation of the receptor by an agonist leads to activation of all intracellular signaling pathways emanating from the receptor via a heterotrimeric G-protein. However, it has recently been demonstrated that GPCRs do not only signal in this simplistic fashion, but rather activate a network of downstream effects comprised of parallel signal transduction pathways. It is now proposed that GPCR ligands that are biased towards induction or blockade of specific signaling pathways will have differentiated physiology compared with unbiased molecules, through selective engagement of a desired subset of signal cascades to deliver a unique set of beneficial effects that can be separable from the undesirable effects generally exhibited by an orthosteric agonist. Such functional selectivity may provide a mechanism to develop new drugs with more precisely targeted effects by activating or inhibiting only the relevant signal transduction pathways. The primary goal of this project is to identify and characterize functionally selective biased ligands for a wide range of GPCRs and to make these available to the scientific community in order to rapidly accelerate and expand the understanding and utility of functional selectivity as a path to new and better medicines. In Specific Aim #1, we will select 12 GPCRs for which a suite of assays will be designed, optimized, and deployed to identify biased ligands against each receptor. The selected GPCRs will be subjected to a robust and validated screening process, through which biased ligands for these GPCRs will be identified. In Specific Aim #2 we will study the selected biased ligands, to improve their bias, potency, and receptor selectivity. This broad scale screening, optimization, and characterization project would dramatically increase the number of potent biased ligands that are available for further study and development, both as research tools and as potential therapeutic candidates. The scientific knowledge resulting from this project has the potential to fundamentally change the way that researchers think about GPCR biology and its role in disease, and to define the way that GPCR ligands themselves are identified, analyzed, and categorized in the future. Functionally selective ligands will be made available to the GPCR community for further investigation of specific GPCR signal transduction pathways. This will dramatically augment understanding and appreciation of functional selectivity by enabling a wide range of in vitro and in vivo experiments. In particular we expect that this project will instigate a rapid advancement in the understanding of functionally selective and nonselective GPCR ligand pharmacology, and enable exploration of the potential for functionally selective molecules to influence pharmacological efficacy, tolerance, and adverse effects of new drug discovery efforts. PUBLIC HEALTH RELEVANCE: Approximately one third of today's FDA-approved drugs act on G-protein coupled receptors (GPCRs), collectively affecting hundreds of millions of people worldwide. In this application, Trevena proposes to demonstrate the broad applicability of functionally selective biased ligands that activate only specific pathways emanating from GPCRs, thus separating the beneficial effects of GPCR activation from the undesirable effects produced by it. The scientific knowledge resulting from this project has the potential to fundamentally change the way that researchers think about GPCR biology and its role in disease, and to redefine the way that GPCR ligands themselves are identified, analyzed, and categorized in the future. This project serves to advance the NIH mission to improve health as it will rapidly accelerate the discovery of a new generation of improved GPCR medicines.